Dendritic cells (DCs) have essential roles in early detection of pathogens and in subsequent activation of both innate and adaptive immune responses. The role of DCs in infection with HIV-1 remains poorly understood, even though these cells are likely to be of critical importance in developing new strategies for protective HIV vaccination. The goal of this proposal is to gain a better fundamental understanding of how HIV-1 interacts with DCs and how this interaction affects the ability of the DCs to coordinate an anti-viral immune response. Even though DCs express cell surface receptors for HIV-1 entry, they are resistant to productive viral replication due to a block that occurs before reverse transcription of the viral genome can be completed. However, DCs have a unique ability to "hand-off" the virus to CD4+ T cells, and thus greatly enhance the infectivity of these cells. HIV-1 can be coaxed into infecting DCs if the cells are first exposed to virus-like particles (VLPs) that deliver the Vpx protein of the related primate lentivirus, SIVmac. Vpx inactivates a dominant restriction for HIV-1 replication in DCs and can therefore be used to determine how the immune response is affected if HIV-1 can productively infect DCs. In preliminary studies, we have found that MDDC infected with HIV-1 (using SIVmac VLPs containing Vpx) up-regulate co-stimulatory molecules involved in the activation of T cells as well as a type I interferon response, suggesting that viral infection triggers an innate immune signaling pathway in these cells. Specific Aim 1 will be to determine how HIV-1 infection of monocyte-derived DC (MDDC) triggers innate immune signaling pathways. Gene expression profiling will be performed on MDDC to provide a bioinformatics guide for the pathways to be investigated, and RNAi approaches will then be employed to identify host cell molecules involved in induction of costimulatory molecules and other innate immunity genes. The stage of HIV-1 replication at which the host innate immune response is activated will be determined by using a combination of HIV replication inhibitors and viral mutants. Specific Aim 2 will examine the consequence of DC infection with HIV-1 on the spread of virus in cultures of DC and T cells and on anti-viral and other anti-microbial T cell responses. The spread of HIV-1 (encoding Vpr or a Vpx-Vpr fusion that permits replication in DC) and the induction of antigen-specific T cell responses will be investigated in co-cultures of MDDC and CD4+ T cells. By comparing infection under conditions that permit or restrict infection of DCs, it will be possible to determine if natural resistance of DC to infection with HIV-1 is beneficial to the host or to the virus. Together, these approaches will provide new tools for studying anti-HIV immune responses and for developing more effective vaccine strategies. PUBLIC HEALTH RELEVANCE: Understanding the nature of the immune response in the HIV-infected individual lies at the heart of our efforts to develop effective prophylactic or therapeutic vaccines for AIDS. Our proposal is to better characterize the interaction of HIV-1 with dendritic cells, which make up the early microbial detection system in the body. Results of these studies may provide new approaches for inducing strong protective immune responses against the virus.